This application relates to low radiocarbon dietary supplements for vertebrates including mammals, fish and fowl, and particularly for humans, which supplements are preferably directed to nucleotides, amino acids, and other DNA and histone protein precursors, and to methods for making such supplements, preferably using byproduct or recycled carbon dioxide “greenhouse gas” emissions from industrial combustion processes involving fossil fuels.
The majority of carbon in the biosphere (the atmosphere, oceans, and surface of the earth where life exists) consists of carbon-12 (about 98.89% natural abundance) or carbon-13 (about 1.11% natural abundance). These are both stable (non-radioactive) isotopes. However, there is a third significant carbon isotope, carbon-14 (also referred to elsewhere and in this document as 14C or radiocarbon), which is radioactive with a half-life of 5730 years and which decays by beta emission to nitrogen-14, the stable form of nitrogen. This transmutation of carbon-14 to nitrogen-14 involves beta particle (electron) decay with an energy of 0.1565 MeV (156,500 eV).
The formation of radioactive carbon-14 in the earth's upper atmosphere was first theorized in the 1930's by Serge Korff, who proposed that cosmic rays striking particles in the upper atmosphere would result in secondary neutrons which would become thermalized and ultimately react with nitrogen-14 atoms to yield carbon-14 and a proton (Currie, 2004). These carbon-14 atoms quickly become oxidized to 14CO2 (radioactive carbon dioxide), which within a few hundred years reach an equilibrium concentration in the atmosphere and oceans with the stable forms of CO2, assuming that the cosmic ray flux is relatively constant. Since virtually all living organisms derive the carbon incorporated into their various structures either directly from CO2 in the atmosphere or oceans via photosynthesis, or directly or indirectly by consuming photosynthetic organisms, during their lifetimes they also will have a percentage of radiocarbon in all of their organic matter that is approximately the same as that of the atmosphere. In the 1940's Willard F. Libby developed a technique to measure these trace levels of radiocarbon in CO2 and in living and dead organic material, and to deduce the age of dead organic material by the corresponding depletion of radiocarbon due to its decay. For this development of the carbon dating technique he was awarded a Nobel Prize (Libby, 1960). Libby experimentally verified that natural background radiocarbon levels of CO2 (referred to herein as the natural abundance concentration of carbon-14) in the biosphere (defined as CO2 in the atmosphere, CO2 dissolved in the oceans, and in organic matter from living organisms all over the earth) had a radioactivity level of about 15.3 dpm/g (disintegrations per minute per gram of carbon). This amount of radioactivity corresponds to a carbon-14 level of just one atom out of every 750 billion total carbon atoms. This natural background radiocarbon level is an approximation, and over time may increase due to additional carbon-14 released into the atmosphere from human or natural nuclear activity, or may decrease by release of additional low radiocarbon into the atmosphere from the burning of fossil fuels. Changes in solar activity may also affect the background radiocarbon levels over time. For the purposes of this disclosure and claims, reference to natural, natural abundance, background or natural background radiocarbon levels, such as “95% below natural radiocarbon levels,” is based on assuming a current natural background level of radiocarbon of about one carbon-14 atom per every 750 billion total carbon atoms. For convenience, natural background levels of radiocarbon may also be expressed in units of percent modern carbon (pMC), where 100% of the current natural background radiocarbon level is equal to 100 pMC. The terms radiocarbon level and radiocarbon concentration are used interchangeably.
Asimov (1955, 1976) was the first to clearly point out that even this trace level of radioactive carbon-14 would result in a significant number of mutations in human DNA. However, he accepted this natural background level of radiocarbon and the resulting mutations as unavoidable. Asimov instead focused attention on preventing any further increase in radiocarbon levels by limiting atmospheric testing of nuclear weapons. Further, Asimov speculated on whether a diet high in C-14 would increase the mutation rate in Drosophila (the fruit fly) or if it would increase tumor formation rates in cancer-prone strains of rats.
Carbon-14 decay occurring in carbon atoms in living organisms may cause damage by several mechanisms. First, a beta decay converts a carbon-14 atom into the chemically dissimilar nitrogen-14 atom. This alone can significantly alter the chemical structure of the affected compound or molecule in which it is present. Second, the emission of the 156,500 eV beta particle (electron) results in a massive recoil of the remaining nitrogen-14 atom, ripping it from whatever compound or molecule of which it was originally a part (also referred to in the art as the “Szilard-Chalmers Effect”). Typical C—C bond enthalpies (bond energies) of 3.59 eV, or C—N bond enthalpies of 3.16 eV, would not be strong enough to retain such a recoiling atom, and it is torn from its original structure. Third, the emitted 156,000 eV beta particle can subsequently ionize thousands of nearby molecules, including water molecules and whatever organic molecules are present. Assuming an average of 35 eV needed to form an ion pair, a single emitted carbon-14 beta particle could produce as many as 4400 ion pairs before dissipating all of its energy. Furthermore, each of the ion pairs produced could potentially be free radicals or other chemically reactive species, which in turn could react with other nearby molecules, including DNA or histone protein residues. These three mechanisms of carbon-14 damage are particularly harmful to the cell when the decaying carbon-14 atom is itself part of the DNA or histones of the chromosomes.
Natural sources of carbon with low radiocarbon levels do exist, primarily in ancient limestone, oil, and coal formations that are not in equilibrium with the carbon of the biosphere. Because half of a sample's carbon-14 atoms will decay every 5730 years, a sample starting out at 100 pMC (that is, at 100% of the current natural abundance level) and that is isolated from the biosphere and has no further carbon-14 added to it will have decayed to lower percentages of carbon (pMC) as illustrated in the following table, Table 1:
TABLE 1DecayRemainingTimeRadiocarbon(years)Carbon (pMC)42595.087090.05,73050.07,57040.09,95530.013,30520.019,04010.024,7705.032,3602.034,7001.538,1001.043,8000.5057,0000.1095,0000.00195,0001.0E−03140,0004.4E−06266,5001.0E−12
Absent contamination from biosphere sources, within 1,000,000 years even a massive coal deposit would be reduced to a level of 0 pMC and would not be expected to contain even a single atom of carbon-14. It should be noted that elimination of natural background carbon-14 from ordinary carbon is also theoretically possible using artificial isotopic separation or enrichment processes. However, such processes are extremely expensive and therefore cost prohibitive for use in producing large quantities of materials for food or nutritional purposes. Furthermore, processes are known for producing enriched carbon-13, typically containing unspecified levels of carbon-14, for use in producing organic compounds for isotopic tracer purposes, for example in drugs or other ingestible components. Such enriched carbon-13 compounds are not necessarily lower in carbon-14, since typical isotopic enrichment processes used to separate the heavier carbon-13 from lighter carbon-12 will typically also enrich the concentration of the still heavier carbon-14, unless special measures are deliberately taken. Furthermore, carbon-13 enriched materials, regardless of their carbon-14 content, would likely not be suitable or safe for use as a component of the human (or mammalian) diet because of potential biochemical or physiological issues involving kinetic isotope effects associated with the highly enriched carbon-13 materials.
Matthews (U.S. Pat. No. 5,471,785) proposed a method to reduce the amount of radiocarbon in organisms to extremely low levels using organic nutrients with radiocarbon levels of at most 1/100th the natural abundance level, which he calculated to be less than 2 atoms of carbon-14 per 1014 atoms of carbon (which can also be expressed as 1.5 pMC when adjusted for the natural abundance level of 1 C14 per 750 billion carbon atoms, as assumed in this disclosure). It is disclosed that achieving such low levels of radiocarbon requires fairly drastic methods. There must be a minimum of contact with the atmosphere throughout the entire process, which would require hermetic enclosures or airlocks and purging to eliminate contamination from sources such as atmospheric CO2. Vacuum heat treatment of solid carbon sources such as coal, and use of shielded underground greenhouses with artificial illumination are also recommended. Miekka (U.S. Pat. No. 5,956,896) proposed a variation on Matthews in which a controlled greenhouse using CO2 virtually free from radiocarbon is used to raise photosynthetic plants for human or animal consumption, as well as low radiocarbon-containing animals and animal products for human consumption. The methods disclosed are directed to a total diet low in carbon-14, in order to lower the carbon-14 level in all organic compounds of the organism, including, e.g., lipids, proteins, sugars, etc.
Additionally, ultra-low carbon-14 raw materials having radiocarbon levels below 1 pMC and as low as 0.0001 pMC (see, e.g., Matthews) may not be readily available. Although one would expect (as did Matthews) that most coal or oil deposits, which are conventionally dated as tens or hundreds of millions of years old, should have near 0% radiocarbon, that is not the case. For whatever reason, whether due to contamination by microorganisms, radioactive byproducts of trace amounts of radioactive substances including uranium, small amounts of de-novo synthesis of carbon-14 in-situ, or other undefined causes, these deposits are typically found with carbon-14 levels that would be expected for deposits as young as 25,000 to 45,000 years, or 5 to 8 half-lives of carbon-14 (Lowe, 1989). Thus, coal radiocarbon levels range from about 3.1% to about 0.4% of natural background levels (about 3.1 pMC to about 0.4 pMC, or 1/30th to 1/250th of the natural radiocarbon abundance found in the biosphere).
Various workers have documented that nucleosides and various related DNA precursors are naturally found in human milk, and have provided methods of supplementing infant formula, humanized milk, and other human nutritional formulas with various DNA precursors. See, for example, Gil et al. (U.S. Pat. Nos. 4,544,559, 5,066,500, 1991), Masor et al. (U.S. Pat. Nos. 5,488,039, 5,492,899, 5,602,109, 5,700,590), Kulkarni et al. (U.S. Pat. No. 5,712,256, 1998), Germano (U.S. Pat. No. 6,503,506), and Gohman et al. (U.S. Pat. No. 6,511,696, U.S. Pat. No. 6,645,543). Humans and other mammals have a variety of active and passive nucleoside transporter proteins which facilitate cellular uptake of these compounds (Kong et al., 2004).
Ordinary nucleotide supplementation of infant formulas is currently available; see, for example, “Similac Advance” and “Similac NeoSure Advance—Infant Formula with Iron” (Ross Nutritionals, Abbott Laboratories). Among other nutrients, these formulas contain added amounts of four basic nucleotides: adenosine 5′-monophosphate, cytidine 5′-monophosphate, disodium guanosine 5′-monophosphate, disodium uridine 5′-monophosphate.
In addition to individual amino acid supplements, typically available over-the-counter, are more complex formulations used for special medical purposes. For example, an infant formula containing, among other nutrients, added amounts of the essential amino acids L-cystine, L-tyrosine and L-tryptophan, is commercially available in the United States as “Similac Alimentum Advance” (Ross Nutritionals, Abbott Laboratories). Also available (Mead Johnson Nutritionals) are a number of special dietary replacement formulas supplemented with selected amino acids for use by infants, children, juveniles and adults suffering from metabolic disorders such as phenylketonuria (PKU), maple syrup urine disease (MSUD), homocystinuria, and glutaric acidemia Type 1. “Phenyl-Free 1” and “Phenyl-Free 2” (for infants, children and adults with Phenylketonuria, or PKU) both include 17 added amino acids (all except for phenylalanine, asparagine, and glutamine). “LMD”(for “Leucine Metabolism Disorders”) is a formula for infants and toddlers that includes among other nutrients 16 added amino acids (all the primary amino acids except for leucine, asparagine, glutamic acid, and glutamine). “BCAD 1” and “BCAD 2” (for MSUD) both include 15 added amino acids (all except for leucine, isoleucine, valine, asparagine, and glutamine). “HCY 1” and “HCY 2” (HCY for homocystinuria) both include 17 added amino acids (all except for methionine, asparagine, and glutamine). “GA” (for Glutaric Acidemia Type 1) includes among other nutrients 15 added amino acids (all the primary amino acids except for lysine, tryptophan, asparagine, glutamic acid, and glutamine). Mead Johnson Nutritionals also makes a special formula (“Phenyl-Free 2 HP”) that is provided for expectant women with maternal PKU. It has a higher proportion of amino acids relative to carbohydrates and fats, thus permitting the patient to consume larger amounts of other foods that are not too high in phenylalanine. Other examples of infant formula supplemented with amino acids include Gohman, et al. (U.S. Pat. No. 6,511,696, 2003; U.S. Pat. No. 6,645,543, 2003) and Schmidl (U.S. Pat. No. 5,719,133).
Greenhouse Gases—Emissions of Carbon Dioxide There continues to be a concern that the significant amounts of carbon dioxide (CO2) being released into the atmosphere may act as a “greenhouse gas” with the potential for causing destabilizing climatic change. The United States Department of Energy (DOE) released figures showing that the worldwide emission of carbon dioxide reached 25,162 million metric tons in 2003, a 37% increase since 1980. This same report disclosed that 98% of the CO2 emissions from the U.S. were from the combustion of fossil fuels such as coal, oil, or natural gas. At least 35% of the CO2 emissions in the U.S. come from stationary combustion sources, such as coal-fired electrical generators or industrial heaters or furnaces. The National Energy and Technology Laboratory of the United States Department of Energy (DOE) (NETL DOE, www.netl.doe.gov) is spending millions of dollars funding numerous carbon sequestration research projects aimed at reducing emissions from such CO2 sources into the atmosphere and thus minimize the greenhouse gas effect. The goal of these projects and other international efforts is to find ways to convert the CO2 into biomatter or other organic forms, even if the resulting material is merely dispersed in the ocean or deposited in abandoned mines as inert solid waste. Current estimated costs of sequestering CO2 from a coal-fired power plant are significant, ranging from $35-$264 per ton. The DOE's goal is to reduce this cost to below $10 per ton. One DOE report (Capture and Sequestration of CO2 From Stationary Combustion Systems by Photosynthesis of Microalgae) describes efforts to use photobioreactors with microalgae to sequester CO2 emissions into high value products including pharmaceuticals, fine chemicals and commodities. However, conversion to products such as low radiocarbon nucleotides or amino acids has never been recognized or suggested. Further improvements in dietary supplements having reduced labels of radiocarbon will be particularly useful.